In this work we will purify the already identified principal antigenic cell surface and corresponding soluble released glycoproteins of clonal PC12 pheochromocytoma and cultured sympathetic neuron cells. Specific antibodies will be prepared against each of these components including the 230 kd NILE glycoprotein, the components with apparent molecular weights of 180 kd, 160 kd, 140 kd, 118 kd, and 55 kd and cytoskeleton-associated 25-30 kd glycoprotein. The localization of each of the antigens on NGF treated and untreated PC12 cells, on sympathetic neurons in culture and on brain cultures will be determined by light and electron microscopic immunohistochemistry. Anatomical distributions in the adrenal medulla, brain, and sympathetic ganglia will also be determined. The effect of the antibodies and antigens on differentiation of PC12 cells in culture will be assessed with respect to cell-substrate adhesion, cell-cell aggregation, neurite formation and organization of neurites into fascicles. Complement-mediated cytotoxicity will be used to select and characterize mutants lacking the surface glycoproteins in an effort to understand the role and function of these macromolecules. The glycoproteins will be covalently linked to substrates and the effect of these substrates on growth and regeneration of neurites will be investigated. A variety of other biochemical and physiological indices of differentiation will also be monitored in these studies. Such experments will be performed both to study the functional role of these glycoproteins and to further the use of the antisera and antigens as experimental reagents. Purified antibodies will be provided to the laboratory of Dr. Ira Black for investigation of the glycoproteins during embryologic development. Finally, a radioimmunoassay for the released glycoproteins will be developed with the aim of immunodiagnosis of neural crest tumors in animals.